This study investigates the structural basis for drug-induced long QT syndrome (LQT), a condition that predisposes individuals to lethal arrhythmias. The HERG K⁺ channel, which is crucial for cardiac repolarization, is often blocked by commonly used medications, leading to acquired LQT. Using alanine-scanning mutagenesis, the researchers identified specific residues in the HERG channel that are critical for high-affinity drug binding, particularly the methanesulfonanilide antiarrhythmic drug MK-499. The binding site, confirmed through homology modeling, is located in the S6 transmembrane domain and the pore helix, specifically at amino acids G648, Y652, and F656. Other structurally unrelated compounds, such as terfenadine and cisapride, also interact with these residues. The aromatic residues Y652 and F656, unique to HERG K⁺ channels, are key to drug binding. The findings suggest that the high-affinity binding site for drugs like MK-499 is likely in the inactivated state of the HERG channel, and the structure-activity relationship can be used to develop tools for drug screening to avoid HERG channel blocking compounds.This study investigates the structural basis for drug-induced long QT syndrome (LQT), a condition that predisposes individuals to lethal arrhythmias. The HERG K⁺ channel, which is crucial for cardiac repolarization, is often blocked by commonly used medications, leading to acquired LQT. Using alanine-scanning mutagenesis, the researchers identified specific residues in the HERG channel that are critical for high-affinity drug binding, particularly the methanesulfonanilide antiarrhythmic drug MK-499. The binding site, confirmed through homology modeling, is located in the S6 transmembrane domain and the pore helix, specifically at amino acids G648, Y652, and F656. Other structurally unrelated compounds, such as terfenadine and cisapride, also interact with these residues. The aromatic residues Y652 and F656, unique to HERG K⁺ channels, are key to drug binding. The findings suggest that the high-affinity binding site for drugs like MK-499 is likely in the inactivated state of the HERG channel, and the structure-activity relationship can be used to develop tools for drug screening to avoid HERG channel blocking compounds.